Our goal is to investigate mechanisms of synaptic transmission mediated by excitatory amino acids. The studies will focus on mammalian neocortical neurons, because of their rich investiture with excitatory amino acid receptors, using in vitro brain slices and primary dissociated cell cultures. State of the art, quantitative, neurophysiological techniques, including single-electrode current- and voltage-clamping, will be used to address basic questions of synaptic organization and function. The overall goal is to better understand the role of various excitatory amino acid receptors in neurotransmission and the processes regulating receptor sensitivity. The specificity, potency and mechanism of action of non-N- methyl-D-aspartate (NMDA) receptor antagonists in the neocortex will be studied. This will be accomplished by using current- and voltage-clamp techniques in the slice preparation and patch-clamp methods on cultured neurons. The role of non-NMDA receptors in synaptic transmission will be studied. We will determine if these receptors affect only excitatory inputs to neurons or if inhibitory inputs, via effects on interneurons, are also altered. The possible unmasking of NMDA-mediated components of synaptic transmission after blockade of non-NMDA receptors will be studied under normal and epileptic conditions. The effects of zinc and glycine, putative NMDA receptor modulators, on synaptic transmission in the neocortex will be studied in brain slice, preparations by examining intracellularly recorded EPSPs and responses to iontophoretically applied excitatory amino acid agonists. Studies on cultured rat neocortical neurons will determine the mechanism underlying the biphasic response, which we have recently described, of these neurons to certain excitatory amino acids The dose dependency of these responses will be examined and interactions between agonists tested. The role of desensitization and intracellular regulatory factors will be evaluated. Whole-cell patch-clamp techniques, in conjunction with rapid perfusion techniques, will be used in these studies on cultured neurons. Additional studies in this preparation will examine the role of calcium entry in transmitter release mediated by excitatory amino acids. Synaptic potentials evoked by application of excitatory amino acid agonists will be studied under normal conditions and after blocking calcium entry. These studies will evaluate whether calcium release from intracellular stores can promote transmitter release.